Hyaluronic acid is an important component of the extracellular matrix of gingivae and its quantity and molecular size appear to be altered under inflammatory conditions. Whether gingival fibroblasts from inflamed tissues synthesize quantities and molecular sizes of hyaluronic acid that differ from normal gingival fibroblasts is not known. To determine this, we isolated fibroblasts from three biopsies each of healthy and chronically inflamed human gingiva and incubated them in the presence of [3H]glucosamine. The release of labeled macromolecules into the medium was approximately 50% greater for the inflamed tissue fibroblasts than for the normal tissue fibroblasts. Of this labeled material, 35% was identified as hyaluronic acid in the medium of inflamed cell cultures, compared to only 25% in normal fibroblast cultures. Sepharose CL-4B chromatography of the labeled material revealed that most of the newly synthesized hyaluronic acid was of large molecular size in inflamed fibroblast cultures. The proportions of [3H]-labeled hyaluronic acid in these peaks varied and indicated that an increase in the amount of large molecular size hyaluronic acid was responsible for the increase in labeled hyaluronic acid noted in the medium of the inflamed tissue fibroblasts. Thus, the decrease in molecular size of hyaluronic acid previously noted in inflamed tissue most likely arises from extracellular factors rather than synthesis of smaller molecular weight species by the fibroblasts. More importantly, however, the differences noted between normal and inflamed gingival fibroblasts persisted over time in culture. This indicates that such differences between the cells may be of a stable and heritable nature.